Marijuana (Cannabis sativa) is the most commonly abused illicit drug by teens and young adults of childbearing age in the USA with significant social and public health implications. While research efforts have begun to characterize the behavioral and neurobiological consequences of direct repeated exposure to cannabis, the possible impact across multiple generations is not known. Preliminary data obtained in our lab suggests that adult rats from parents with adolescent exposure to D-tetrahydrocannabinol (THC; primary psychoactive component of cannabis) exhibit reduced motivation and a depression-like phenotype, as well as anxiety-related behaviors that develop in response to their tendency for increased heroin intake. In addition, both male and female F1 offspring show decreased striatal expression of genes that encode several components of the dopamine, opioid and glutamate neurotransmitter systems. Female F1 offspring tended to have more significant disturbances, emphasizing the importance of gender that could potentially be relevant to sex differences seen in psychiatric disorders. The goal of this proposal is to characterize the whole population of mRNAs in the nucleus accumbens (NAc) of F1 offspring from parents with a history of THC exposure by transcriptome sequencing, as well as to study whole genome DNA methylation and chromatin modifications to identify the epigenetic milieu underlying the cross- generational disturbances in this mesolimbic brain region. The molecular studies will be combined with behavioral tests to assess reward-, depression- and anxiety-like behaviors in F1 males and females. We will address the causality between gene expression impairments and specific behavioral phenotypes using in vivo gene manipulations in the NAc. In order to determine whether there is persistent transgenerational transmission, phenotypic and epigenetic marks shown to be impaired in the F1 generation will be followed into the F2 progeny. Overall, these studies will provide fundamental mechanistic insights into the cross-generational impact of cannabis exposure that could guide the development of therapeutic interventions for the spectrum of behavioral phenotypes associated with vulnerability to addiction and psychiatric disorders.